Verena Ruckstuhl, Thermo Fisher Scientific, Langenselbold, Germany and
Julie Foster, Ph.D., Thermo Fisher Scientific, Asheville, NC
Appli c a ti on N ote
Determination of Bacteria Retention in the
Thermo Scientific Barnstead Smart2Pure
Water Purification System
Key Words
Water purification, lab water, ultrapure water, bacteria, 0.2 micron filter,
Thermo Scientific Barnstead, Smart2Pure
Abstract
The bacteria retention of a Thermo Scientific™
Barnstead™ Smart2Pure™ water purification system
was evaluated using the membrane filtration method
based on the European Pharmacopoeia method described
in Chapter 2.6.12.1
Introduction
Bacteria are single celled organisms that can be found
nearly everywhere in a busy laboratory. Although many
of these bacteria are harmless to a person’s health, they
can create unwelcome variability in an experiment.
Because of their abundance and ability to be easily
transferred, precautions such as sterile technique were
employed. Using bacteria-free water during steps such as
sample preparation, system rinsing, or buffer preparation
is an easy method of reducing the chance of bacterial
contamination.
Water purification systems are a reliable source for
bacteria-free water. On average, bacteria such as
Escherichia coli (E. coli), which are widely used in the
laboratory, have an length of about 2 µm, and an average
diameter of 0.5 µm.2 A 0.2 µm absolute membrane filter
at the end of the system is used to remove any particles
or bacteria that are larger than the pore size of the filter3.
Proper maintenance of the water system, including filter
replacement as specified in the manual, helps to ensure
the water remains bacteria-free.
Ultrapure water from a Barnstead Smart2Pure water
system was analyzed for the presence of aerobic bacteria.
The Barnstead Smart2Pure 3 system was chosen from
the family of Smart2Pure systems, which also includes
Smart2Pure UV and UF models. All of these systems have
the same feed water requirements, basic flow path and all
dispense water through a 0.2 µM final filter. The systems
are fed by tap water and utilize two purification steps
to create 18.2 megohm lab water. The first step purifies
the feed water using a reverse osmosis (RO) membrane
to remove the majority of impurities in the water. The
second step of purification utilizes an ultrapure cartridge
to remove any ions in the water. Additionally, an optional
ultraviolet (UV) lamp and/or an ultrafilter (UF) can be
added to the system configuration.
Methods
Bacteria retention of the Smart2Pure system using
tap water as feed water
The bacterial content of tap water was determined after
disinfecting the water outlet with 70% ethanol and
rinsing with 1 L water. Afterwards three 1 L sample
volumes were collected and tested for bacterial growth
using the membrane filtration technique.1 Three different
volume samples were generated from the three 1 L
samples: a 1 L, 10 mL, and a 1 mL sample. These
three samples were filtered through a 0.2 µm cellulose
nitrate (CN) membrane. The membrane was afterwards
transferred aseptically to a R2A-Agar and incubated at
35°C for 5 days in a Thermo Scientific™ Heratherm™
compact microbiological incubator (model IMC18).
The agar plates with sample volumes of the membrane
filtration (1 L and 10 ml) showed after 5 days incubation
at 35°C too many colonies to count. Only the 1 ml
sample volumes gave results that could be quantitated.
After installing and rinsing of the Smart2Pure 3 system
according to the Smart2Pure water purification system
operating instructions, the system was ready for
sampling. 0.2 L of water from the system was used to
rinse the 0.2 µm filter of the Smart2Pure 3 system and
then 1 L samples were collected in sterile flasks.
Figure 2. Set up of the filtering of the 1 L sample, using
a Thermo Scientific Nalgene analytical test filter funnel,
through the 0.2 µm cellulose nitrate (CN) membrane.
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ITALIAN
Tap water
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Tap water
Smart2Pure 3 system
Appli c a ti on N ote
SPANISH
Figure 1. Diagram of a Smart2Pure water system with a 6 L internal reservoir.
Rusia +7 812 703 42 15
Suiza +41 44 454 12 22
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Analysis
of
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the
samples
Otrosopaises
Analysis f t+49
he 6184 90 6000
Analysis f to
he o +33 2 2803 2180
samples sample Sterile tube
GenPure Pro Sterile flask Tutti gli (altri) marchi sono di proprietà di Thermo Fisher Scientific Inc. e delle sue filiali. Specifiche,
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condizioni e tariffe possono subire variazioni. Non tutti i prodotti sono disponibili in tutti i paesi.
Per maggiori
Test tube dettagli consultare il rivenditore locale.
Smart2Pure 3 The 1 L sample was then filtered through a 0.2 µm
Australia +61 39757
Indiathe
+91 tap
22 6716
2200line (1 L and 10 ml)Svizzera +41 44 454cellulose
12 22
Table4300
1. Bacteria count from
water
nitrate (CN) membrane, using a Thermo
Austria +43 1 801 40 0
Italia +32 02 95059 552
UK/Irlanda +44 870 609 9203 ™
Nalgene™ analytical test filter funnel, as shown
Belgio +32 53 73 42 41
Olanda +31 76 579 55 55
USA/Canada +1 866Scientific
984 3766
Cina +800 810 5118 or
Nuova Zelanda +64 9 980 6700
in Figure 2. The membrane was afterwards transferred
Tap Water Sample
(10del
ml)
(1 L)Asiatiche +852 2885 4613
+400 650 5118
Europa
Nord/CIS Smart2Pure System Sample
Altre Nazioni
aseptically
a R2A-Agar and incubated at 35°C for
Francia +33 2 2803 2180
+358 9 329 10200
Nazioni non in elenco
+49 6184 90to
6000
Germania
+49 6184 90 6000
Russia +7 812 703 42 15
o +33 2 2803 2180 5 days in a Heratherm compact microbiological incubator
1st Run
Giappone +81 3 5826 1616
Spagna/Portogallo +34 93 223 09 18
(model IMC18). The Smart2Pure 3 system was disinfected
with the disinfection routine according to operating
GERMAN
instructions, and the pretreatment cartridge, ultrapure
thermoscientific.com
polisher cartridge and 0.2 µm filter were replaced with
new ones to retest the system.
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Results
Australien +61 39757 4300
2nd +32
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Belgien
China +800 810 5118 or
+400 650 5118
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0800 1 536 376
Deutschland international
+49 6184 90 6000
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FRENCH
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After 5 days, any bacteria colonies found on the
703 42 15
Russland +7 812
R2A-Agar
Spanien/Portugal +34
93 223 09 18plates were counted and the amount of
Schweiz +41 44 454bacteria
12 22
per ml in the water were calculated. The data
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is
summarized
in Table 1.
USA/Kanada +1 866 984 3766
Andere asiatische Länder
+852 2885 4613
Conclusion
Andere Länder +49 6184 90 6000
The
Smart2Pure
system,
oder +33 2 2803 2180
which was directly connected to
tap water, was able to remove the bacterial load of the
tap water down to < 0.01 CFU/ml.
References
1.European Directorate for the Quality of Medicines &
Pharmacopoeia 6.8,
Non-sterile Products:
Microbial Enumeration Tests, 2010
Thermo Fisher Scientific Inc. Tous
Tapdroits
Water
Les (autres) marques déposéesSmart2Pure
System Water
© 2013
réservés.
sont des marques commerciales
ou déposées de Thermo Fisher
HealthCare (EDQM): European
Scientific Inc et de ses filiales. Les caractéristiques, conditions et tarifs sont susceptibles d’être modifiés. Tous les produits ne sont pas disponibles dans tous les
Bacteria
Count
n.a.
<0.01
Microbiological Examination of
pays. Pour tout renseignement, veuillez vous adresser à votre distributeur local.
(CFU/ml)
Allemagne +49 6184 90 6000
France +33 2 2803 2180
Australie
+61
39757 4300
Inde +91to22evaluate
6716 2200
n.a. not
applicable,
the colonies were too numerous
Autriche +43 1 801 40 0
Italie +32 02 95059 552
Belgique +32 53 73 42 41
Japon +81 3 5826 1616
Chine +800 810 5118 or
Nouvelle-Zélande +64 9 980 6700
+400 650 5118
Pays Bas +31 76 579 55 55
Espagne/Portugal +34 93 223 09 18
Pays nordiques/baltes/CEI
+358 9 329 10200
Russie +7 812 703 42 15
Suisse +41 44 454 12
22
2.Thermo
Scientific Waterbook. 2014. Thermo
UK/Irlande +44 870 609 9203
Scientific.
Retrieved 12 June, 2014 from
USA/Canada +1 866 984 3766
http://www.thermoscientific.com/purewater
Autres pays d’Asie +852 2885 4613
Autres Pays +49 6184 90 6000
3.Operational
ou +33 2 2803 2180
Instructions for the Thermo Scientific
Barnstead Smart2Pure water purification system.
Thermo Fisher Scientific, Niederelbert, Germany.
thermoscientific.com/purewater
thermoscientific.com
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